1. Development of Five Dual-Color, Double-Fusion Fluorescence in Situ Hybridization Assays for the Detection of Common MLL Translocation Partners 
Jeannette G. Keefe, William R. Sukov, Ryan A. Knudson, Lai P. Nguyen, Cynthia Williamson, Jason P. Sinnwell, Rhett P. Ketterling 
The Journal of Molecular Diagnostics 
Volume 12, Issue 4, Pages (July 2010)
DOI: /jmoldx
Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

2. Figure 1
Probe design schematics for MLL, AFF1, MLLT4, MLLT3, ELL, and MLLT1 as used in the dual-color D-FISH probe assays. A: Probe schematic for the five BACs labeled in Spectrum Green, RP11-832A4, RP11-215H18, RP11-770J1, RP11-861M13, and CTC-89C10, spanning the 11q23 MLL region. B: Probe schematic for the four BACs labeled in Spectrum Orange, RP11-397E7, RP11-168E22, RP11-476C8, and RP11-529H2, spanning the 4q21 AFF1(AF4) region. C: Probe schematic for the five BACs labeled in Spectrum Orange, RP11-931J21, RP3-431P23, RP11-471L1, RP3-470B24, and RP11-164L23, spanning the 6q27 MLLT4(AF6) region. D: Probe schematic for the four BACs labeled in Spectrum Orange, RP11-15P13, RP11-336O12, RP11-73E6, and RP11-4E23, spanning the 9p22 MLLT3(AF9) region. E: Probe schematic for the four BACs labeled in Spectrum Orange, RP11-282P1, CTD-3137H5, CTD-2516F17, and CTD-2643B8, spanning the 19p13.1 ELL region. F: Probe schematic for the five BACs labeled in Spectrum Orange, CTC-232P5, CTB-66B24, CTC-503J8, RP11-114A7, and RP11-30F17, spanning the 19p13.3 MLLT1(ENL) region. Note: figures are not drawn to scale.
The Journal of Molecular Diagnostics  , DOI: ( /jmoldx )
Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

3. Figure 2
General strategy for the MLL pairing partner D-FISH assay. A: Each of the five assays uses the MLL probe labeled in green and a second probe labeled in red for one of the translocation partner genes (AFF1, MLLT4, MLLT3, ELL, or MLLT1). B: Ideograms of chromosome 4 and 11 with red (R) and green (G) ovals representing the 4q21 AFF1 and 11q23 MLL probe regions, respectively. Normal chromosome four homologues display a 2R signal pattern and normal chromosome 11 homologues display a 2G signal pattern. Reciprocal translocation of chromosomes 4 and 11 resulting in disruption of AFF1 and MLL causes splitting of one of the R and G signals, respectively. Subsequent juxtaposition of the split AFF1 and MLL signals results in two sets of paired R and G signals, observed as two yellow F signals. The normal chromosomes 4 and 11 remain as separate R and G signals. An overall 1R1G2F FISH signal pattern is observed.
The Journal of Molecular Diagnostics  , DOI: ( /jmoldx )
Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

4. Figure 3
Representative interphase FISH signal patterns observed with the MLL/partner D-FISH assays. A: Nonneoplastic nucleus with a normal 2R2G signal pattern. B: Nonneoplastic nucleus with coincidental R and G signal overlap resulting in a 1R1G1F signal pattern. C: Neoplastic nucleus with a 1R1G2F signal pattern representing a balanced translocation resulting in two MLL/partner fusion signals. D: Neoplastic nucleus demonstrating a 2R2G1F signal pattern representing a complex translocation. E: Neoplastic nucleus demonstrating a 1R2G1F signal pattern indicating an unbalanced MLL/partner translocation with an associated deletion of the partner DNA at one of the translocation junctions. F: Neoplastic nucleus demonstrating a 2R1G1F signal pattern indicating an unbalanced MLL/partner translocation with an associated deletion of the MLL DNA at one of the translocation junctions. G: Neoplastic nucleus with a 1R1G3F signal pattern due to gain of an additional copy of one of the derivative chromosomes. H: Neoplastic nucleus with a 1R3G1F signal pattern indicating an unbalanced translocation and MLL duplication/trisomy of chromosome 11. I: Neoplastic nucleus with a 2R1G2F signal pattern indicating a balanced translocation and an additional copy of the partner chromosome. J and K: Abnormal signal patterns lacking a fusion (3R1G and 2R3G) are observed as a result of translocations that disrupt only one of the two genes covered by the probe set. The 3R1G signal pattern also denotes a deletion of 1 MLL signal. Arrows denote yellow fusion signals.
The Journal of Molecular Diagnostics  , DOI: ( /jmoldx )
Copyright © 2010 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions